Sasmal D, Guhathakurta B, Ghosh A N, Pal C R, Datta A
National Institute of Cholera and Enteric Diseases, Calcutta, India.
FEMS Microbiol Lett. 1992 Nov 1;77(1-3):217-24. doi: 10.1016/0378-1097(92)90159-l.
An N-acetyl-D-glucosamine-specific cell associated hemagglutinin (HA) was isolated and purified from a strain of Vibrio cholerae 01 by chitin affinity chromatography followed by separation on Bio Gel P-150. A single stained protein band of 47 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was observed with the purified HA. HA-antisera produced a single precipitin band against the purified HA in an immunodiffusion test without exhibiting any reactivity towards purified lipopolysaccharide (LPS). Purified HA, used as solid-phase antigen in an enzyme-linked immunosorbent assay (ELISA), reacted strongly with HA-antisera but cross-reacted negligibly with antisera raised against purified LPS. Hemagglutinating activity of the purified HA was highly sensitive to N-acetyl-D-glucosamine. The immunogold-labelling method using HA-antisera confirmed the location of the HA on the surface of the bacterial cells. The HA-antisera reacted with a protein component of the homologous outer membrane preparation. A significant inhibition was observed in the adhesive capability of the V. cholerae 01 strain to isolated rabbit intestinal epithelial cells (RIEC) in vitro when the later were pre-treated with the purified HA.
通过几丁质亲和层析，随后在生物凝胶P - 150上进行分离，从一株霍乱弧菌O1中分离并纯化出一种N - 乙酰 - D - 葡萄糖胺特异性细胞相关血凝素（HA）。在十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳（SDS - PAGE）中，纯化后的HA呈现出一条47 kDa的单一染色蛋白带。在免疫扩散试验中，HA抗血清针对纯化后的HA产生了一条单一沉淀带，而对纯化的脂多糖（LPS）未表现出任何反应性。在酶联免疫吸附测定（ELISA）中用作固相抗原的纯化HA，与HA抗血清强烈反应，但与针对纯化LPS产生的抗血清交叉反应可忽略不计。纯化HA的血凝活性对N - 乙酰 - D - 葡萄糖胺高度敏感。使用HA抗血清的免疫金标记方法证实了HA在细菌细胞表面的定位。HA抗血清与同源外膜制剂的一种蛋白质成分发生反应。当用纯化的HA对兔离体肠上皮细胞（RIEC）进行预处理时，观察到霍乱弧菌O1菌株对其体外黏附能力有显著抑制。
